This invention relates to PrPSc-specific antibodies and to peptides used for their generation. These antibodies are suitable for detecting PrPSc in a sample, and for purifying PrPSc. Additionally, the invention relates to diagnostic aids for the detection of PrPSc, pharmaceuticals that contain or mimic PrPSc-specific conformational epitopes, and methods for prion decontamination.
Prions are infectious agents that are associated with neurodegenerative syndromes characterized by spongiform change (e.g., microcavitation of the brain, usually predominant in gray matter), neuronal cell loss, astrocytic proliferation disproportionate to neuronal loss, and accumulation of an abnormal amyloidogenic protein, sometimes in discrete plaques in the brain. It is possible that neurodegeneration in prion diseases shares certain underlying mechanisms with other more common neurodegenerative syndromes such as Alzheimer's Disease, amyotrophic lateral sclerosis, and Parkinson's disease.
The agents that transmit these diseases differ markedly from viruses and viroids in that no chemical or physical evidence for a nucleic acid component has been reproducibly detected in infectious materials (Prusiner, Science, 216: 136–144, 1982). A major step in the study of prions and the diseases that they cause was the discovery and purification of a protein designated prion protein (PrP) (Bolton et al., Science 218:1309–11, 1982; Prusiner et al., Biochemistry, 21:6942–50, 1982; McKinley et al., Cell, 35:57–62, 1983). When purified using proteinase K digestion, a 27–30 kD protease-resistant protein was discovered in scrapie-affected hamster brain, and was termed PrP 27–30, later found to be a fragment of PrPSc (Bolton, Science, 218:1309–1311, 1982).
According to the prion hypothesis, prion infectivity resides in PrPSc. PrPSc is at least strongly associated with infectivity and appears to be a reliable surrogate for prion infection. PrPSc is a conformational variant of a host-encoded cellular protein designated PrPC (Oesch et al., Cell, 40:735–746, 1985), which is a glycosylphosphatidylinositol (GPI)-linked cell surface protein with a molecular mass of 33–35 kD.
PrPC has been isolated from normal brain, and has been found to be protease-sensitive and not associated with scrapie disease-producing activity (Bendheim et al., Ciba Found. Symp. 164–177, 988). According to the prion theory, PrPC converts into PrPSc autocatalytically (Prusiner, Proc. Natl. Acad. Sci, USA 95:13363–83, 1998). More recently, it was reported that PrPC can be converted to a protease-resistant form in vitro by PrPSc (Kocisko et al., Nature, 370:471–473, 1994). PrPC is an evolutionarily conserved membrane protein for which the actual biological or physiological function is unknown. Mice devoid of PrPC are viable and show no obvious signs of neurological and physical impairment (Bueler et al., Nature, 356:577–582, 1992). Additionally, these mice are not susceptible to prion infection, underscoring the central importance of PrP in the replication of infectivity (Bueler et al., Cell, 73:1339–1347, 1993; Prusiner et al., Proc. Natl. Acad. Sci. USA, 90:10608–10612, 1993). Targeted investigations of PrP knockout mice revealed impaired synaptic function (Collinge et al., Nature, 370:295–297, 1994) and altered sleep regulation (Tobler et al., J. Neurosci., 17:1869–79). Moreover, PrPC has been shown to modify T cell activation induced by concanavalin A stimulation (Cashman et al., Cell 61:185–192, 1990), indicating a functional role for the protein.
The prion diseases are a group of rapidly progressive, fatal, and untreatable neurodegenerative syndromes. Human prion diseases include Creutzfeldt-Jakob disease (CJD), which has sporadic, iatrogenic, and familial forms; and variant CJD (“vCJD”), likely derived from the consumption of cattle tissues contaminated with the agent of bovine spongiform encephalopathy (reviewed in Cashman, Can. Med. Assoc. J. 157:1381–5, 1997). CJD has been accidentally transmitted between humans by contaminated cadaveric pituitary hormones, dura mater transplantation, neurosurgical instrumentation, and corneal transplantation (Brown et al., Lancet 340:24–7, 1992). The potential risk of transmitting CJD through blood and blood products is of worldwide concern. Moreover, scrapie in sheep and goats is a common and economically important prion-related disease in North America, as is bovine spongiform encephalopathy (BSE) in Great Britain. According to Britain's Ministry of Agriculture, Fisheries and Food, more than 4,347,380 cattle have been destroyed, because they were deemed old enough to conceivably harbor the disease agent. The Ministry of Agriculture has estimated that the total cost of the epidemic will reach $7.13 billion by 2002. More than 173,000 bovines from all over Britain have been confirmed to be infected, and hundreds of thousands more might have entered the food supply undetected.
Additionally, the United States and Canada have now implemented a blood donor deferral for individuals who resided in the UK during the early and peak years of the BSE epidemic. Such a restriction has been adopted as a precaution against the risk of transmitting a vCJD, which to date has afflicted over 60 Britons since 1996. The Canadian Blood Services estimates that 120,000 of its 600,000 active donors, or 22%, have visited Britain since 1980 (Montreal Gazette, May 6, 1999). Many new donors have been recruited to replace the loss, raising several concerns. One such concern is that the blood of new donors is not as safe since it has only been screened once for illnesses such as hepatitis and human immunodeficiency virus.
Accordingly, a need exists for diagnostic methods suitable for mass screening of prion infected blood or tissues. The availability of antibodies that distinguish PrPC from PrPSc would therefore be of great value in development of a test for prion infection. Furthermore, a need exists for therapeutic agents that prevent and/or treat prion diseases.